PROJECT SUMMARY Alzheimer's disease is characterized by accumulation of insoluble forms of amyloid-? (A?) in the extracellular space and within vessel walls, aggregation of the microtubule protein tau in intracellular neurofibrillary tangles and neuroinflammation - contributing to neuronal damage and clinical dementia. A critical barrier in treating Alzheimer's disease is the years to decades-long lag between initiation of disease and clinical diagnosis, when reversal of brain pathology and recovery of neurons may at best, slow cognitive decline. Thus, it is essential to identify the pathological processes in early disease to characterize biomarkers for diagnosis and find interventional targets that will halt disease prior to symptoms and signs of dementia. An early phenotype of Alzheimer's is olfactory dysfunction. Using a novel, targeted RNA sequencing strategy (BioSpyder TempO- Seq), we found that compared to controls, viral/amyloid processing pathways are activated in OBs and olfactory tracts dissected from formalin-fixed, paraffin-embedded (FFPE) tissue of individuals with familial and sporadic Alzheimer's disease (FAD and SAD). The discovery of viral infection in the OB and OT in Alzheimer's opens a potentially exciting, new avenue of research that links 2 disparate, underexplored areas of Alzheimer's disease pathogenesis: (1) olfactory dysfunction in early Alzheimer's disease that is characterized by the loss of sense of smell, by A? deposition in the glomeruli of the OB, and by stage- dependent deregulation of specific pathways including neuroinflammation in olfactory bulb, and; (2) the association of herpesviruses with Alzheimer's disease, including the increased frequency of herpes simpex virus type 1 (HSV-1) in brains from Alzheimer's disease patients and HSV-1-induced acceleration of amyloid deposition in mice and human neural cultures. Taken together, we hypothesize that productive HSV-1 infection within the olfactory epithelium results in inflammatory activation of glomeruli in the OB of AD patients, overall producing a neuroinflammatory environment that accelerates olfactory dysfunction and progression of AD. To test this hypothesis, we will: Aim 1. Use intranasal inoculation with HSV1 to study microglial responses and formation of amyloid in the OB, and olfactory dysfunction in 5xFAD and controls, and; Aim 2. Compare the gene expression profiles of HSV-1-infected OB and OT from the 2 cohorts of animals described above with that of previously analyzed FAD and SAD OB and OT using the BioSpyder TempO-Seq RNAseq assay. The experiments proposed here are within the scope of the parent grant DC000566 that investigates through combined behavioral, two photon imaging and electrophysiological recording olfactory function in mice and are significant since they will identify pathogenic mechanisms that occur in early AD prior to the onset of clinical dementia that can provide early biomarkers for diagnosis and therapeutic targets to slow or halt progression of AD.